The Mentored Clinical Scientist Development Award (K08) is intended to provide support for clinicians committed to research to facilitate their development into independent investigators. The K08 award is well suited for this candidate, who has completed clinical and research training, but requires more experience and publications before starting an independent research career. Dr. Hornstra will work in the laboratory of Dr. Steven Shapiro, under the guidance of Drs. Shapiro and Welgus. The sponsor, co- sponsor and collaborator at Washington University are well know for their expertise in the field of matrix biology. This unique environment will greatly enrich the candidate's potential to study the lysyl oxidases, important extracellular matrix cross-linking enzymes. Funding this candidates KO8 proposal will allow the generation of significant preliminary data and publications for future R29 or R01 research proposals. The objective of the research project is to investigate the role of lysyl oxidases in extracellular matrix development and maintenance. Lysyl oxidases are primordial enzymes which function to cross-link collagen and elastin in the presence of molecular oxygen, producing the strong and flexible ligaments and tendons of large animal bodies. In the human and now mouse, two lysyl oxidase genes have been identified: lysyl oxidase-1 and lysyl oxidase-2. In the mouse, only lysyl oxidase-1 (mLO-1) has been characterized, but my preliminary work has identified the murine lysyl oxidase-2 gene (mLO-2). These two genes exhibit differential expression in human and mouse cell lines and tissues. In many adult mouse tissues, LO-2 mRNA expression is readily detectable but LO-1 mRNA expression is undetectable via Northern Analysis. However, in fetal skin and lung fibroblast cell lines, LO-1 mRNA expression is slightly greater than LO-2, but LO-2 expression is detectable. These data suggest differential mechanisms of gene regulation and potentially cell-specific or tissue- specific expression during normal matrix development and maintenance. Lysyl oxidases are intimately involved with normal extracellular matrix development and turnover, but if abnormally regulated, may contribute to a variety of development and fibrotic disorders. The long term goal of this proposal is to delineate the role of the lysyl oxidase genes in the extracellular matrix of connective tissues during normal development, and in the future during pathological fibrosis. To accomplish this, we propose to generate lines of mice deficient in either mLO-2 and mLO-1 genes, examine the phenotypes of these deficient mice, and examine the differential expression and regulation of the LO genes. Our hypothesis is that either mLO-2 or mLO-1 deficient mice will have a specific phenotype due to the temporal and spatial differential expression of the LO genes. The phenotypes will result from he specific and independent functions of either LO, and this phenotype will be defined in the "knockout" models. These specific functions of either mLO-2 or mLO-1 could not be identified with the use of pharmacological inhibitors, which are non-specific. In the unlikely event that no significant phenotype is observed or can not be induced upon injury challenge, a double "knockout" with both LOs would be constructed in the future.